PROJECT SUMMARY Genetic testing has made it possible to identify individuals with germline predisposition to cancer, e.g., carriers of ATM, BRCA1, TP53 mutations. In addition to an increased risk of developing primary cancer, germline mutation carriers may develop post-treatment-related malignancies (i.e., therapy-related leukemias), which develop as new primary malignancies, rather than as relapse of the original tumors. I have recently shown that germline mutation carriers may be at an increased risk of clonal hematopoiesis of indeterminate potential (CHIP). CHIP is defined as somatic expanded blood cell clones in individuals without other hematologic disease. CHIP is identified by gene variants found on next-generation DNA sequencing assays that are present at very low frequency. These variants do not represent the germline, hematologic malignancy, post-zygotic mosaicism, or circulating tumor burden. CHIP rates in healthy populations have been estimated to be 2% in those under the age of 60, and increase with age and/or exposure to chemotherapy. The clinical significance of CHIP lies in the increased risks for cardiovascular disease, primary and secondary leukemias, other hematologic malignancies, and in general for all-cause mortality. I have recently gained data supporting the notion that ATM mutations represent a novel germline predisposing factor to CHIP. In fact, I have shown that heterozygous ATM germline mutation carriers: 1) are relatively frequent in the general population of healthy individuals (1 in 263 individuals of European ancestry) and women with familial breast cancer (1.5 in 100 individuals); 2) have an apparent increased rate of CHIP as compared to non-ATM mutation carriers; and 3) may have unique molecular CHIP signatures. Therefore, ATM germline mutations are common and may predispose to CHIP. ATM regulates the DNA damage response to double-strand DNA breaks through its kinase activity. Due to ATM haplo-insufficiency and DNA-repair defect predisposition, I postulate that ATM germline mutation carriers may not have the proper mechanisms of DNA repair and therefore cannot prevent CHIP, particularly after cytotoxic therapy (chemotherapy or radiation). Guided by my preliminary data, I will test the hypothesis that ATM germline mutations are associated with and predispose hematopoietic cells to develop CHIP. I will test my hypothesis through the following Specific Aims: 1) to identify the clinical and biologic factors associated with ATM germline mutations and CHIP and lay the foundation for prospective population-based studies, 2) to explore chromosomal instability and dynamics of CHIP in a prospective cohort of ATM germline mutation carriers, and 3) determine if CHIP can be recapitulated in a mouse model and predict clinical malignant evolution in ATM mutation carriers. In summary, CHIP may pose a notable risk for ATM mutation carriers and determination of how it arises and what drives its progression is necessary to counsel, screen and manage this patient population appropriately.